In an attempt to get a better understanding of the interactions between Beta-adrenergic receptors (BAR), adenylate cyclase and membrane components, monoclonal antibodies have been raised using BAR solubilized from frog erythrocyte plasma membrane as the antigen. The immunization was performed both in vivo and in vitro. ELISA and indirect immunoprecipitation of BAR labeled with 125I-iodohydroxylbenzylpindolol were used as the screeining for antibody production. One of the monoclonal antibodies has been massively produced and extensively characterized. This antibody (with a chain composition of Igg-1-K) partially precipitated BAR solubilized from membranes of frog erythrocytes, frog heart and C6 glioma cells. When the antigen was fractionated Sephadex G-150 column chromatography, two BAR binding peaks were detected; only the binding peak with higher molecular weight could be immunoprecipitated. Immunoblotting of an isoelectrofocusing gel of the antigen reveals that the antibody was bound to a single spot with a PIof about 6.2. Immunoprecipitation of total erythrocyte proteins labeled with 125I indicated that this antibody interacted with a protein of approximately 43,000 daltons. Internalized BAR (which was free of adenylate cyclase and guanine nucleotide binding protein) failed to be immunoprecipitated. This antibody affected biphasically basal and isoproterenol-sensitive adenylate cyclase activity in isolated plasma membranes of frog erythrocytes. At low concentration of the immunoglobulin, the cyclase activity was activited, whereas at higher concentration the activity was inhibited by the antibody. These characteristics suggest that this antibody is against a membrane component (such as guanine nucleotide binding protein) which interacts with both BAR and adenylate cyclase. This monoclonal antibody may be a useful tool for the study of the regulation of BAR function.